The synthesized polymer was characterized by size exclusion chromatography, proton nuclear magnetic resonance and attenuated total reflection-Fourier transform infrared spectroscopies. The main-chain photo- and acid-induced degradation of the copolymer in dimethylsulfoxide and water, respectively, was verified by UV-vis spectroscopy at light intensities as low as 0.1 mW cm−2 at λ = 254 nm. Next, a model anticancer drug, doxorubicin (DOX), was chemically linked to the polymer chain end(s) via acylhydrazone bond(s), resulting in amphiphilic PEG-alt-adipic acid-DOX  polymer–drug conjugates. The conjugates were self-assembled in water to form spherical nanoparticles, as evidenced by scanning and transmission electron microscopies. The irradiation of the self-assembled PEG-alt-AA-DOX conjugates with UV light and the decrease of the solution pH resulted in the disruption of the assemblies due to the photolysis and acidolysis of the acylhydrazone bonds, and the release of the therapeutic cargo. Light-mediated polymer degradation has attracted considerable attention in various applications, including photo-patterning, tissue engineering and photo-triggered drug delivery. In this study, we report the synthesis and characterization of a new, linear, main-chain photo- and acid-degradable copolymer based on acylhydrazone linkages. The polymer was synthesized via a step-growth copolymerization of adipic acid dihydrazide with a bifunctional poly(ethylene glycol) bearing benzaldehyde end-groups, under mild acidic conditions, to afford a hydrophilic PEG- Stimuli-degradable polymers (SDPs) have gained increasing attention due of their wide range of potential applications in various fields, including nano- and bio-technology as drug carriers and actuators, and in bio-patterning, etc ,2. Recen−2) compared to other polymeric materials [Among the various chemical and physical stimuli  employed to trigger the degradation of the polymer chains, light-degradable polymers have attracted considerable interest. Light, as an external stimulus, offers unique advantages such as spatiotemporal control, as well as the tunability of the irradiation wavelength and intensity . So far,aterials ,8. More aterials .SDPs have been extensively explored as nanocarriers for the controlled and/or programmable delivery of drugs and other active biomolecules, exhibiting very promising results. Polymer micelles, polymersomes, nanogels and polymer–drug conjugates are typical nanostructures employed in drug delivery applications ,15,16,17Linear, main-chain polyhydrazones and polyacylhydrazones belong to a polymer family that has barely been studied in the literature because of their low solubility in both polar and non-polar solvent media ,26. Hydrald), under mild acidic conditions, to afford a hydrophilic PEG-alt-adipic acid  alternating copolymer  and water, was investigated. Copolymers with a hydrazide end-group were prepared using an excess of the adipic acid dihydrazide comonomer during the polymerization. Next, the hydrophobic model anticancer drug, doxorubicin (DOX), was linked to the hydrazide end-groups of the polymer via acylhydrazone linkages, to produce amphiphilic PEG-alt-adipic acid-DOX  drug conjugates   were obtained from Sigma Aldrich  and the doxorubicin hydrochloride salt  was obtained from LC Laboratories . The diethyl ether, petroleum ether, n-hexane (96%) and dichloromethane (DCM) were purchased from Scharlau . The DCM was dried over calcium hydride. The DMSO (99.9%) was obtained from Fisher Chemical  and the ethanol (≥98%) from Honeywell . Milli-Q water, with a resistivity of 18.2 MΩ·cm at 298 K, was obtained from a Millipore apparatus, and was used for the preparation of all of the samples.The 4-carboxybenzaldehyde (98%) and triethylamine  were purchased from Alfa Aesar . The 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide , 4-(dimethylamino)pyridine , adipic acid dihydrazide (≥98%) and poly(ethylene glycol)  and EDC  were dissolved in 20 mL anhydrous DCM, and were stirred at 40 °C for 30 min, followed by the addition of DMAP  and HO–PEG–OH . The reaction was stirred at 40 °C for 24 h. Next, the product was precipitated in diethyl ether, washed 10 times with petroleum ether and dried overnight under vacuum at room temperature (RT). The dried product was dissolved in chloroform and filtered through a 0.45 μm PTFE filter to remove the unreacted 4-carboxybenzaldehyde. Finally, it was precipitated again in diethyl ether and dried overnight under vacuum to obtain PEGald  was added to the solution, followed by the addition of 2–3 drops of 0.1 N HCl. The reaction was left under stirring for 24 h at 25 °C. The next day, the solvent was evaporated using a rotary evaporator, and the reaction was dried under vacuum at RT. The white solid was dissolved in chloroform, and was purified by fractional precipitation in hexane. The final copolymer was stored under a N2 atmosphere at 4 °C until use.Adipic acid dihydrazide  was dissolved in 2 mL milli-Q water. The monomer solution was placed in a sonication bath until its complete dissolution. Next, PEGAdipic acid dihydrazide  was dissolved in 4 mL ethanol. The solution was first sonicated for 10 min and then stirred for 10 min in a water bath at 40 °C until its complete dissolution. Next, a solution of 4-carboxybenzaldehyde  in 2 mL ethanol was prepared. The two solutions were mixed and a white solid precipitated instantly. The reaction was stirred for another 3 h before filtering off the white solid product and washing it with a large amount of ethanol and water. The final product was dried overnight under reduced pressure at RT.alt-AA copolymer  was added to the DOX solution, and the reaction mixture was stirred for 20 h before being transferred to a dialysis membrane with a molecular weight cut-off of 3500 Daltons. The mixture was dialyzed against DMSO to remove the unreacted DOX molecules. The conjugated DOX molecules were quantified by fluorescence spectroscopy using a standard calibration curve of DOX in DMSO, and the drug loading was calculated using Equation (1).First, DOX∙HCl  was dissolved in DMSO (2.5 mL), and TEA (3 μL) was added to neutralize the HCl. After 20 h stirring at RT, hydrophobic DOX was obtained. Next, the PEG-alt-AA-DOX conjugate (5 mg mL−1) in 5 mL DMSO was prepared. Next, 15 mL water (pH 7.4) was added to the solution under vigorous stirring at a rate of 0.1 mL min−1 using a syringe pump. Afterwards, the solution was dialyzed against water using a dialysis membrane with a molecular weight cut-off of 3500 Daltons to remove the organic solvent. The final solution was filtered through a 0.45 μm pore size cellulose acetate syringe filter, and was kept at 4 °C until use.A solution of the PEG-Mn) and the molecular weight distribution (Mw/Mn) of the polymers. THF (containing 2 v/v% triethylamine) was used as the eluent at a flow rate of 1 mL min−1. The calibration curve was based on six narrow poly(methyl methacrylate) (PMMA) standards with molecular weights ranging from 625 to 138,000 g mol−1. In order to prepare the samples for SEC, the polymers were dissolved in THF at a concentration of 15 mg mL−1; the polymer solutions were then filtered through 0.45 μm pore size PTFE syringe filters and injected into the system. 1H Nuclear Magnetic Resonance (1H NMR) spectra were recorded on two Brucker 300 MHz and 500 MHz spectrometers  using CDCl3 or DMSO-d6 as the solvent. Attenuated total reflection-Fourier transform infrared (ATR-FTIR) spectra were recorded on a Nicolet 6700 spectrometer . For each spectrum, 64 scans were collected in the 500–4000 cm−1 range. The ultraviolet–visible (UV-vis) absorption spectra were recorded on a Shimadzu UV-2600 spectrophotometer  in the wavelength range of 200–600 nm. The samples were prepared by the dissolution of the polymers or polymer–DOX conjugates in DMSO or H2O (pH 7.4) at a concentration of 10−3 mg mL−1. A Malvern Zetasizer Nano ZS instrument  equipped with a He–Ne laser (λ = 633nm), at a 90° scattering angle, was used for the dynamic light scattering (DLS) measurements. Before each measurement, the sample was filtered through a 0.45 μm pore size hydrophilic, cellulose acetate syringe filter. The measurements were conducted at 25 °C. The size and the morphology of the self-assembled structures were characterized by field emission-scanning electron microscopy (FE-SEM) using a JEOL JSM-7000F microscope , and by transmission electron microscopy (TEM) using a JEOL JEM-2100 microscope . The samples were prepared by the deposition of a drop of a dilute polymer solution on a glass substrate for SEM or a carbon coated copper grid for TEM, and were dried overnight at 25 °C. The fluorescence spectra were recorded on a Lumina Fluorescence Spectrometer (Thermo Scientific)  equipped with a 150 W CW Xenon-arc lamp. The excitation and emission slits were set at 10 nm and 20 nm, respectively. The fluorescence spectra of the DOX and the PEG-alt-AA-DOX conjugate were recorded from 500 nm to 900 nm upon excitation at 488 nm, whereas the respective spectrum of the copolymer was recorded upon excitation at 302 nm. All of the samples were measured at room temperature.A Waters size exclusion chromatography (SEC) instrument comprising a Waters 515 isocratic HPLC pump, two Mixed-D and Mixed-E (Polymer labs) columns  detector  were used for the determination of the molecular weight , the PEG-alt-AA alternating copolymer (λ = 302 nm) or the PEG-alt-AA-DOX conjugate (λ = 297 nm), before and after irradiation for the different time intervals (t), respectively.All of the photoirradiation experiments were performed using a UV lamp at 254 nm and a power density of 0.1 mW cmalt-AA alternating copolymer solution in water was prepared; its pH was adjusted at 5.2 using HCl, and the solution was kept under stirring at 37 °C. The hydrolysis of the polymer was monitored by measuring the absorption of the polymer solution at predesigned time intervals by UV-vis spectroscopy.A 0.2 mg/mL PEG-alt-AA-DOX conjugate  at pH 7.4, 5.2 and 2.0 were prepared; each solution was transferred to a dialysis bag with a molecular weight cut-off of 3500 Daltons, and was immersed in 10 mL H2O at pH 7.4, 5.2 and 2.0, respectively, under gentle stirring (100 rpm) at 37 °C. Next, the samples were irradiated at 254 nm for 5 min at predesigned time intervals. After each irradiation cycle, the solutions were stirred for 15 min, and then the aqueous media were collected and replaced with 10 mL fresh medium of the respective pH value. An aqueous solution of the self-assembled PEG-alt-AA-DOX conjugate  at pH 2.0 was also prepared as a control sample, and the medium was collected and replaced as described above at the predesigned time intervals without prior irradiation. The collected samples were dried, first using the rotary evaporator and then under vacuum for 20 h. The dry samples were dissolved in DMSO (2.5 mL) and their fluorescence spectra were recorded. The concentration of DOX in each sample was determined using a standard calibration curve of DOX in DMSO. The DOX percentage released was calculated using the following equation:Ct and t−C1 are the concentrations of DOX at two successive irradiation time intervals, and totalC is the initial concentration of DOX in the self-assembled conjugate.Aqueous solutions of the self-assembled PEG-Mn = 4000 gr mol−1), which is hydrophilic, non-toxic and FDA approved. Moreover, it is well known for its stealth properties, and has been used extensively in drug delivery [ald was characterized by 1H NMR and ATR-FTIR spectroscopies and SEC. The 1H NMR spectrum of PEGald (δ 3.67 ppm (H1), which was assigned to the methylene protons of the monomer repeat units of PEG, and two peaks at δ 3.87 ppm and δ 4.53 ppm (H2 and H3) assigned to the methylene protons next to the newly formed ester bonds, verifying the successful derivatization of the polymer end-groups. In addition, the presence of a peak at δ 10.13 ppm, which corresponds to the aldehyde protons (Ha), and the peaks at δ 8.0 and 8.24 ppm, attributed to the aromatic protons (Hb and Hc) of the benzaldehyde group, confirmed the presence of the aromatic aldehyde functionalities at the two polymer chain ends. The ATR–FTIR spectrum of the product (−1) attributed to the aldehyde and ester groups introduced at the polymer chain ends and supported the NMR results. Finally, the SEC traces of the PEG macromonomer , the two comonomers—PEGald and adipic acid dihydrazide—were reacted at a 1:1.2 molar ratio under mildly acidic conditions. The SEC analysis of the polyacylhydrazone polymer prepared , and confirmed the SEC results discussed above. It should be noted that the absence of the peak of the aldehyde proton of PEGald at δ 10.13 ppm suggests that the quantitative reaction of PEGald with the dihydrazide comonomer was in excess in the reaction. ald macromonomer and the PEG-alt-AA alternating copolymer. The characteristic vibration bands at 2869 cm−1, 1694 cm−1 and 1095 cm−1—assigned to the N–H, C=O and C–O stretching vibrations, respectively,—were observed in the spectrum of the alternating copolymer. More importantly though, the presence of the characteristic vibration band of the acylhydrazone bond at 1600 cm−1 in the spectrum of the copolymer, which is absent in the ATR–FTIR spectra of the two precursor comonomers, verified the successful synthesis of the polymeric product.For the synthesis of the alternating polyacylhydrazone copolymers  and its degradation profile was monitored by UV-vis spectroscopy. Before the irradiation, the diacylhydrazone molecule exhibited a broad absorption band centered at ~302 nm  compared to other photodegradable polymers reported in the literature [The light-mediated degradation of the acylhydrazone-based copolymer was further verified by UV-vis spectroscopy. omonomer b. The raomonomer d, albeitterature ,42.alt-AA copolymer prepared in this work, a solution of the polymer in H2O was first irradiated for 2 h at 254 nm to cleave the acylhydrazone bonds, and was afterwards left under stirring for 24 h in the dark. The UV-vis absorption spectra of the copolymer after irradiation and following 24 h in the dark were compared  and ester (δ 4.53 ppm) protons changed and shifted to lower ppm, whereas two new peaks appeared at δ 2.2 ppm and δ 9.8 ppm, which were attributed to the formation of acetaldehyde as a by-product of the photodegradation reaction.As discussed above, poly(acyl)hydrazones have been studied in the literature as dynamic covalent polymers, and have been proposed for the development of self-healing materials and adaptable networks due to the reversible nature of the (acyl)hydrazone bonds, which can be cleaved and reformed upon the application of a stimulus, such as acidic media or temperature ,43,44. Icompared , verifyicompared . In ordealt-AA alternating copolymer was used for the synthesis of the PEG-alt-AA-DOX drug conjugates. For the conjugation of DOX at the ends of the polyacylhydrazone chains, the copolymer and DOX were mixed at a 1:2 mole ratio in DMSO and were reacted for 24 h, followed by dialysis against DMSO to eliminate the unreacted DOX molecules. alt-AA-DOX conjugate in DMSO, in which the polymer peak at 302 nm is clearly observed, whereas the inset shows the magnified spectrum in the 340–700 nm range, where the absorption peak of DOX is evident, indicating the successful conjugation of the drug at the polymer chain ends. alt-AA-DOX conjugate in DMSO. As expected, the copolymer does not emit light in the 500–800 nm range, whereas DOX and the PEG-alt-AA-DOX conjugate both emit at 600 nm, verifying the conjugation of the drug at the polymer chain ends. The drug loading of DOX on the copolymer chains was quantified from the fluorescent intensity of the PEG-alt-AA-DOX conjugate using a calibration curve of DOX in DMSO  in water. We envisaged that the conjugation of the hydrophobic DOX drug moieties at one or both ends of the hydrophilic PEG-d by SEM a, and a d by SEM b. The hyd by SEM , in goodalt-AA-DOX nanoparticles and the release of DOX, upon the cleavage of the acylhydrazone bonds with UV light, was investigated by SEM, TEM and DLS. alt-AA-DOX conjugates in water (pH 7.4) following irradiation with UV light for 2 h. The absence of the PEG-alt-AA-DOX assemblies after irradiation is evident, indicating the main-chain cleavage of the polyacylhydrazone backbone. Smaller nanoparticles, 25–30 nm in size, were observed in the SEM image after light irradiation, which were attributed to aggregates formed by the released hydrophobic DOX molecules. The results were also confirmed by TEM  attributed to the drug aggregates. Finally, the disruption of the self-assembled PEG-alt-AA-DOX nanostructures, upon exposure to UV light, was verified by DLS . As discussed above, the characteristic absorption band of the polymer at ~302 nm decreased with the irradiation time, while a new band at ~260 nm emerged, indicating the main-chain cleavage of the polymer chains and the formation of the precursor PEG macromonomer. A similar degradation process was found for the PEG-alt-AA-DOX assemblies upon the irradiation of a polymer solution adjusted to pH 5.2  during the irradiation; the dialysate was collected after each irradiation cycle, and its DOX content was analyzed by fluorescence spectroscopy. In parallel, another PEG-alt-AA-DOX solution at neutral pH 7.4 was irradiated with UV light, and the release of the cargo in the dialysate was followed by fluorescence spectroscopy. As shown in alt-AA-DOX assemblies, an aqueous PEG-alt-AA-DOX solution at pH 2.0 was irradiated with UV light for 2 h. Indeed, a faster drug release profile was found , at which a synergistic photolysis and acidolysis of the prodrug leads to faster release kinetics. Therefore, the proposed drug delivery system could be used for the effective photo-triggered drug release at specific sites/tissues, in which the pH ranges from neutral  to acidic (cancer tissues), providing an additional level of control over the drug release profile. Further studies are underway to investigate the in vitro effectiveness of the proposed polymer–drug conjugates towards cancer cells.Finally, the effect of light and a low solution pH on the disruption of the PEG-o pH 5.2 b, whilsto pH 5.2 c, suggesalt-AA—in DMSO and water, upon irradiation at 254 nm (0.1 mW cm−2), was studied, and the cleavage of the hydrazone bonds along the polymer main-chain to produce the two precursor comonomers was verified at very low irradiation doses (720 mJ/cm−1). Next, amphiphilic PEG-alt-AA-DOX conjugates were prepared by the condensation of DOX molecules at the polymer chain end(s), and the conjugates were self-assembled in water to form spherical nanoparticles. At neutral and slightly acidic pH values (pH 5.2), the PEG-alt-AA-DOX assemblies were disrupted, and the conjugated drug was released at shorter irradiation times and a low total irradiation dose, whereas the drug-release kinetics were accelerated upon the synergistic effect of light irradiation and a low solution pH (2.0), denoting the remotely controlled drug release nanosystem. We envisage that such photo- and acid-degradable polymers could be used for the conjugation of both hydrophilic and hydrophobic drug molecules, vitamins or other bioactive molecules bearing an aldehyde or ketone group, and are thus attractive for the development of diverse drug carriers. Current studies are underway to investigate the effectiveness of the polymer–drug conjugates towards cancer cells, and to red-shift the irradiation wavelength required for the photodegradation of the polymer acylhydrazone bonds and therefore improve their potential in the biomedical field.In summary, a new, main-chain photo- and acid-degradable, amphiphilic polymer–drug conjugate, based on a hydrophilic polyacylhydrazone chain and a hydrophobic model anticancer drug, DOX, was proposed. The alternating polyacylhydrazone copolymer was synthesized via a step-growth polymerization reaction of adipic acid dihydrazide with dibenzaldehyde-terminated PEG. The photodegradation profile of the copolymer—PEG-


The strategy was based on a simple ionic crosslinking reaction between the carboxyl groups present in the ionomer and zinc oxide (ZnO), where the formation of reversible Zn2+ salt bondings exhibits the self-healing behavior. The heterogeneous blending of EPDM and ionomers was also used to investigate the thermal and mechanical properties of the TPVs. The experimental findings were further supported by the surface morphology of the fracture surfaces viewed using microscopy. The self-healing behavior of the TPVs has been identified by scratch resistance testing, where the EPDM-g-MAH TPVs showed excellent healing efficiency of the scratch surface. Therefore, this work provides an efficient approach to fabricate new ionically cross-linked thermoplastic vulcanizates with excellent mechanical and self-repairing properties for the skins of automotive interior door trims and instrument panel applications.The development of smart elastomeric materials with inherent self-repairing abilities after mechanical damage has important technological and scientific implications, particularly in regard to the durability and life cycle of rubber products. The interest in self-healing materials for automotive applications is rapidly growing along with the increasing importance of vehicle scratch quality and quantity. The creation of a reversible network by noncovalent ionic cross-linking in elastomer/rubber blends is an effective approach to generate the self-healing phenomenon, with reprocessing and recycling properties. In this work, thermoplastic vulcanizates (TPVs) were prepared using ethylene–propylene–diene (EPDM) polymers and high-acid-containing thermoplastic ionomers. Along with the general EPDM, maleic anhydride grafted EPDM (EPDM- Rubbers are extensively used in various fields, especially in vehicle tires and different automotive parts. Due to increased road accidents, scratches on automotive parts occur quite often, decreasing the durability and life cycle of the products. Self-healable rubber is expected to contribute to the advanced manufacturing of different automobile parts ,2. The cUntil now, self-healing research has been predominantly focused on polymeric systems that heal through chemical means. The presence of ionic associations or crosslinking in rubbers and ionomers has also been noted as a powerful approach for designing self-healing rubbers ,20,21. XIn the present work, ionomer/EPDM composites were prepared through the dynamic vulcanization process. Along with the general EPDM rubber, maleic anhydride-grafted-EPDM rubber was also used. The thermal and mechanical properties, along with the compression set of the composites, were investigated. The surface morphology of the vulcanizates was also explored. The self-healing behavior of thermoplastic vulcanizates has also been identified by a scratch-resistance test. The developed material is expected to be applied to the skins of automotive door trims and for instrument panel applications. From the viewpoint of sustainable development, this efficient strategy of constructing reversible dynamic crosslinks in a solid state shows a good prospect for the rational design of EPDM rubbers for perceived applications in the automobile industry. g-MAH), grade KEPA1150, in pellet form, were purchased from Kumho Petrochemical Ltd., Daejeon, Korea. Ionomer (HA60D) grade High acid 60D, partially neutralized with zinc, was supplied by WAPS Co., Ltd., Busan, Korea. The carboxylic acid (acrylic acid) content of the ionomer is 20% and the degree of neutralization is 50%. Reagent grade zinc oxide and stearic acid were obtained from Daejung Chemicals & Metals Co., Ltd., Seoul, Korea. The 2-2′-Dithiobis (benzothiazole) (MBTS), tetramethylthiuram disulfide (TMTD), and sulfur were procured from Sigma-Aldrich, Seoul, Korea . All the compounding chemicals were used as received without further purification. In this research, ethylene–propylene–diene rubber (EPDM) grade KEP570P with an ethylene content of 70 wt%, Mooney Viscosity ML (1 + 4) at 125 °C of 53, and ethylene–propylene–diene monomer-grafted maleic anhydride (EPDM-g-MAH/HA60D relative concentrations (wt%) used for this study are 60/40 and 70/30. First, EPDM and Ionomer (HA60D)  were premixed at 130 °C and 60 rpm for 8 min. Then, the first set of compounding ingredients was added and mixed for another 2 min. Finally, sulfur was added to the mixture and further mixed for 8 min. EPDM-g-MAH/HA60D blends  were also prepared using the same mixing procedure. The compounds were kept at room temperature for 24 h before conducting the cure characteristics assessment. TPVs of different compositions were prepared using the melt mixing technique in an internal mixer following the method in After the melt mixing, all the blends were compression molded and vulcanized at 170 °C for 10 min to obtain 2 ± 0.1 mm thick plain sheets.2 atmosphere at 10 °C/min heating rate from −70 to 300 °C for all TPV blends and pure components. The glass transition temperature (gT) and melting temperature (mT) were taken as the inflection points from the heating curve to identify the heterogeneous phase structure of the TPVs compounds. Differential Scanning Calorimetry (DSC): Thermal analysis was carried out in a TA Instruments DSC Q20 , under NThermogravimetric (TGA) Analysis: The thermal degradation characteristics of pristine compounds and TPV blends were obtained using a TA Instruments DSC Q50   under nitrogen atmosphere from room temperature to 600 °C at a heating rate of 10 °C/min.Mechanical Properties: The tensile properties of the dumbbell-shaped specimens were measured in a Myungji Tech Tensometer 2000  at a crosshead speed of 500 mm/min. The thickness of the specimens was kept as 2 ± 0.1 mm. The stress-strain curves of five samples were recorded and the average values are reported. Hardness and Compression Set: The hardness was evaluated using a Shore A durometer  following ASTM D2240. The compression set (CS) % was measured with a Myungji Tech compression set  using cylindrical samples with a diameter of 29 ± 0.5 mm and a thickness of 12.5 ± 0.5 mm, according to ASTM D395. The samples were compressed using two parallel plates to a linear deformation of 25% at 70 °C for 22 h.Microscopy Analysis: To elucidate the morphology of the TPVs, field-emission scanning electron microscopy  was carried out at an accelerating voltage of 20 kV. A cryo-fractured sample from a tensile specimen was used for analysis. The samples were mounted on aluminum stubs with carbon tape and then sputtered with gold coating for observation.Self-Healing Scratch Test: A self-healing test was performed using a scratch hardness tester . The scratches were created by applying a 0.50 mm diameter round ball-type tungsten carbide (WC) tip on the TPVs surface at a constant load (10 N) and speed (40 mm/s); then the sample was placed in a convection oven maintained at 89 °C for 24 h. The test method was followed according to BMW GS97034-9 for leather-type structured plastic materials for car interiors. The change of scratch width was monitored using the laser confocal microscope  technique. The self-healing efficiency was calculated as follows:g-MAH, and HA60D, were dynamically vulcanized to fabricate self-healing TPVs. The EPDM-g-MAH polymer was used to investigate compatibility issues in the blends. It was observed that the EPDM-g-MAH showed better compatibility with the ionomer compared to the pristine EPDM. We also considered the effect of sulfur content on the mechanical properties of the EPDM-g-MAH/HA60D composites. A number of blends, consisting of different contents of EPDM, EPDM-2+ ions pairs self-aggregate to form an ionic cross-linked network. The schematic of the reaction and ionic cross-links formation in the thermoplastic vulcanizates is shown in The strategy is based on the simple reaction between the acid groups of ionomer and zinc oxide in the TPVs, where the Zng-MAH mechanical properties were measured. By using the ionomer (HAD60) with EPDM in the TPVs, the tensile strength was increased from 4.9 MPa to 7.8 MPa. With EPDM-g-MAH/ HAD60 (60/40 wt%), the tensile strength increased from 6.0 MPa to 8.8 MPa. As expected, with the decrease in the ionomer content for the 70/30 wt% blend, there was a drop in tensile value. This significant improvement of tensile properties represents the formation of ionic cross-linking with the ionomer phase during the curing process. A higher TS value in the case of EPDM-g-MAH/ HAD60 TPVs might also indicate an improved interaction between the rubber phases and ionomer phase [The introduction of ionic crosslinking into a polymer blend is expected to improve the mechanical properties of the compounds. The interaction of the ionomer with ZnO in the composites was confirmed by the rise in mechanical strength of the TPVs. er phase . g-MAH exhibited a high elongation of 1170 and 600%, respectively. With the addition of the ionomer, the elongation at break decreases to 220% for EPDM/HAD60 (60/40) and to 250 and 270% for EPDM-g-MAH/HAD60 (60/40 and 70/30) blends.In addition, the pristine EPDM and EPDM-g-MAH compounds, the compression set is higher, whereas with the addition of the ionomer in the corresponding vulcanizates, the compression set decreases. This could be due to the presence of higher crosslinks in the multilayered vulcanizates that resist deformation, thus improving the compression set property. To determine the effect of ionic crosslinks and ionomer loading in the TPVs, the compression set property was measured at 70 °C for 22 h. The compression set property provides an insight into the degree of ionic interaction with the acid group of the ionomer, as well as the ability of thermoplastic vulcanizates to retain their elastic properties after the prolonged action of compressive stresses. g-MAH compounds was observed at about 54 and 66 Shore A. It could be clearly seen that the addition of HA60D to the compounds with same amount of filler loading yielded a significant effect on the hardness value. For the 60/40 EPDM and EPDM-g-MAH compounds, the hardness value increased to 73 and 84 Shore A, respectively. Higher crosslink density in the vulcanizates is responsible for higher hardness. Moreover, with the lowering of the EPDM-g-MAH/HAD60 (70/30) ionomer loading, the hardness value decreased.g-MAH provides the maximum effect of compatibilization with the ionomer, thus enhancing the mechanical properties of the vulcanizates. The mechanical strength results were used to represent the compatibility and the quality of the produced EPDM-g-MAH/HAD60 thermoplastic vulcanizates. In addition, the effect of the sulfur content  on the mechanical properties of the TPVs was investigated. g-MAH/HAD60 (60/40) TPVs. The TS increased to 12 MPa with 2.0 phr sulfur content and decreased to 10.4 MPa with the 3.0 phr sulfur compound. In addition, the vulcanizates showed increased hardness with increasing sulfur loading, as shown in Therefore, the presence of polar groups in EPDM-g-MAH, HAD60, and the vulcanizates. The glass transition temperature (gT) is an indirect representation of the heterogeneous nature of the polymeric vulcanizates. The results are summarized in gT of the neat EPDM and EPDM-g-MAH compounds were observed at −47 and −55 °C, respectively. In contrast, the melting point (mT) of the HA60D and EPDM-g-MAH compounds were found at +82 and 162 °C, approximately. The EPDM/HA60D (60/40) vulcanizate showed two distinct peaks (gT and mT) corresponding to the two constituents for the compositions of the blend. For the EPDM-g-MAH/HA60D vulcanizates, a single gT value and two different melting peaks were obtained: one well below room temperature, accordant with the rubber phase, and the other at about 80 °C, in line with the ionomeric phase, thus manifesting the heterogeneous phase structure of the composites. DSC was used to investigate the thermal behavior of the self-healable thermoplastic vulcanizates. g-MAH/HA60D vulcanizates showed higher thermal stability compared to the neat compounds. In addition, when comparing the effect of the EPDM and EPDM-g-MAH, it was observed that the maleic anhydride grafted EPDM provides a slightly better thermal stability to the vulcanizates.g-MAH compounds and the EPDM/HA60D and EPDM-g-MAH/HA60D vulcanizates. From the images (g-MAH/HA60D compound (g-MAH in the ionomer (HA60D) matrix. This is attributed to phase inversion during melt mixing, the common morphology observed in the dynamically cured blend of a thermoplastic and an elastomer.To provide an intuitive observation of the inside structure of the thermoplastic vulcanizates, SEM imaging of the cryogenically fractured surface was performed. e images a,b, we se images c, we seecompound d, the mog-MAH/HA60D and EPDM/HA60D vulcanizates taken before and after thermal aging. An initial scratch with the width of around 300~500 μm gradually decreases to around 40–80 μm after thermal exposure at 89 °C and then becomes unclear after 24 h. The EPDM-g-MAH/HA60D mixtures exhibit a high scratch recovery rate above 87% within 24 h, regardless of the compositions  and (70/30) thermoplastic vulcanizates showed good shape recovery of about 88% after surface-scratch healing. On the other hand, due to the uneven surface structure of the EPDM/ionomer vulcanizates, it was difficult to measure their scratch recovery performance. Finally, it can be concluded that the formation of ionic crosslinks in thermoplastic vulcanizates opens up new opportunities for the development of high performance and smart elastomeric materials. The present study hopes to open new avenues for the sustainable application of cross-linked rubber composites showing recyclable and healable properties, where the effects of the dynamic and thermoreversible nature of ionic crosslinks dominates the permanent covalent bonds.This works reports the preparation of self-healing thermoplastic vulcanizates based on a EPDM polymer and a thermoplastic ionomer (60/40 and 70/30 wt%) by melt-mixing. The study also found that the use of EPDM-


P = 0.003). 7 anastomotic complications were reported  of which five (83.3%) in the cRY were anastomotic stenosis versus none in the pRY group (P = 0.044).  There were no significant differences between the cRY group and pRY group regarding age, sex, BMI, neoadjuvant therapy, preoperative comorbidities, history of laparotomy, ASA score, tumor location, pathological stage, total operative time, incision length, blood loss, time-to-first flatus, time-to-first soft diet, and postoperative hospital stays. The proportions of patients who received a 21 mm stapler were higher in the cRY group (7/44) than that in the pRY group (0/68) ( The application of pant-shaped anastomosis for esophagojejunostomy after LTG is a safe and feasible procedure and has an advantage when the jejunum diameter is small. With the advent and increasing experience of minimally invasive techniques and technology, laparoscopic total gastrectomy (LTG) has gradually gained popularity because of its minimal invasiveness compared with open total gastrectomy (OTG). However, postoperative anastomotic complications such as leakage and stenosis sometimes still occur. A recent meta-analysis showed that the incidence of anastomotic complications was slightly higher in LTG than in OTG . EsophagIn the current status of LTG, most surgeons still prefer extracorporeal anastomosis through a minilaparotomy  rather than total laparoscopic intracorporeal anastomosis, which is more difficult. End-to-side Roux-en-Y esophagojejunostomy by using a circular stapler is widely accepted as a standard reconstruction method, with low rates of anastomotic leakage. However, although seldom reported, the difficulty of stapler insertion sometimes occurs during operation. An obvious difficulty is the insertion and fixation of the anvil head through a narrow window. By using the orally inserted anvil (OrVil™) or reverse puncture device technique, this problem has been solved to some extent , 6. AnotFrom January 2016 to December 2020, our team (led by L. Shi) performed 112 LATG with D2 lymph node dissection for middle-upper-third gastric cancer. Of these, 44 received a conventional Roux-en-Y end-to-side esophagojejunostomy, and the remaining 68 received pant-shaped Roux-en-Y esophagojejunostomy . All patUnder general anesthesia, patients lay in the supine position, with the head higher than the feet (forming a 15-degree incline). The surgeon stood on the left side of the patient, the first assistant was on the right, and the camera operator stood between the legs of the patient. A 12 mm trocar was inserted through an infraumbilical incision. After pneumoperitoneum was achieved, two right 5 mm assistant ports and two left operator ports (5 mm lower and 12 mm upper) were inserted under laparoscopy. After inspection of the peritoneal cavity, the left lobe of the liver was lifted using a suture retraction technique . The stoOnce the stomach was completely mobilized, a 6-10 cm vertical midline incision was made in the upper abdomen, and the fully mobilized stomach was pulled out of the abdominal cavity. Then, the stomach was transected 1-2 cm proximal to the gastroesophageal junction. Choice of the circular stapler size, 25 mm or 21 mm, is based upon the esophageal and jejunal diameter. Commonly, a 25 mm anvil head  was inserted into the esophageal stump under direct vision, and a purse-string suture was tied around its central rod. Next, the jejunum was divided 20 cm distal to the Treitz ligament. The Roux-en-Y jejunojejunostomy was created using a continuous 3-0 Vicryl suture , approximately 50 cm distal to the site of future esophagojejunostomy extracorporeally.The main unit of the 25 mm circular stapler  is inserted through the open jejunal end of the Roux limb with the central rod penetrating the antimesenteric wall 5 cm from the end of the loop. The anvil and circular stapler body were connected, and an end-to-side esophagojejunostomy was conducted under direct view. The instrument is then withdrawn and the open end of the jejunal loop is closed with a linear stapler .First, the proximal end of the Roux limb was folded on itself in a form similar to an “inverted J,” 10 cm long. A 2 cm enterotomy was made at the top corner of the folded jejunum loop with a purse-string suture around it Figures . Then, tth postoperative day. All participants were postoperatively followed up at the outpatient clinic at 1, 3, 6, and 12 months after discharge. Stenosis at the site of the esophagojejunostomy was suspected when patients reported dysphagia, and endoscopic examination was ordered. Stenosis was diagnosed during endoscopic examination when the endoscope could not pass through the anastomosis. All the patients without symptoms were examined 1 year after surgery for direct observation of the anastomosis site.A nasojejunal tube is placed routinely and early enteric nutrition started within 24-48 hours postoperatively. Sips of water were allowed on the 2nd postoperative day, and a fluid diet was usually permitted on the 5th. All patients underwent routine water-soluble contrast swallow X-ray before restarting oral nutrition. The patients are usually discharged on the 11-14t-test or Mann–Whitney U test was used for comparing continuous data, and the chi-square test or Fisher's exact test was adopted for comparing categorical data. P values <0.05 indicated statistical significance.Statistical analysis was performed with SPSS 26.0 software . Continuous data are presented as mean ± standard deviation (SD) or median (range), and categorical data are presented as number (%). Student's A total of 112 patients underwent successful LATG. Of these, 44 patients  received a conventional Roux-en-Y end-to-side esophagojejunostomy (cRY group), and the remaining 68 patients  underwent pant-shaped Roux-en-Y esophagojejunostomy (pRY group). The detailed information of these patients is shown in P < 0.05). The seven cases choosing a 21 mm circular stapler for an esophagojejunostomy were all due to anastomotic trouble caused by the narrow diameter of the jejunum.The surgical outcomes and postoperative complications within 12 months after surgery are summarized in P = 0.028) of which four (66.7%) in the cRY were anastomotic stenosis versus none in the pRY group . Our initial purpose was to investigate the feasibility and efficacy of pant-shaped anastomosis in esophagojejunostomy in LATG; so, we only applied a charge of 60 mm length linear stapler cartridge to construct a small jejunal pouch. Thus, the reservoir effect is small, and it was not in the scope of this study.By December 2020, our team had performed 68 LATGs with pant-shaped Roux-en-Y anastomosis. Compared with our previous 44 LATG with conventional Roux-en-Y anastomosis, the present study showed that pant-shaped Roux-en-Y was feasible and safe in terms of operative time, intraoperative blood loss, and overall postoperative complications. As to the incidence of anastomotic complications, the pRY group was lower than that in the cRY group. In the pRY group, all esophagojejunostomies were successfully performed with a 25 mm circular stapler. In the cRY group, a certain proportion (7/68) of patients could only receive 21 mm stapler. Our results showed that the rate of anastomotic complications, particularly stenosis, is relatively high in cases where 21 mm staples were used . In Zuiki's study, stenosis occurred in 3 (60%) of 5 patients when the 21 mm end-to-end anastomosis stapler, which is close to our results. We think that this may contribute to the high incidence of anastomotic complications in the cRY group. First, using a 21 mm stapler itself leads to a smaller anastomotic diameter. Second, repeated attempts to insert a mismatched stapler, sometimes with violent operation, inadvertently affect the integrity of the intestinal mucosa at the anastomosis site.π-shaped method, the rates of anastomotic leakage still exist [Although several methods for side-to-side anastomosis using linear stapler have been reported to reduce anastomotic stenosis, such as functional end-to-end anastomosis, the overlap method, and the ll exist –22. Morell exist .The application of pant-shaped anastomosis for digestive tract reconstruction after LATG proved to be safe and feasible. When the diameter of the jejunum is small, this method is simple and has an advantage over conventional Roux-en-Y esophagojejunostomy. This study provides a new method for laparoscopic gastrointestinal reconstruction.


The control adhesive was formulated with BisGMA, TEGDMA, HEMA, and the binary system CQ-EDAB as a photo-initiator system. For the experimental adhesives, the EDAB was completely replaced for the TTME or the TTAC monomers. The adhesives formulated with TTME or TTAC monomers achieved double bond conversion values close to 75%. Regarding the polymerization rate, materials formulated with TTME or TTAC achieved lower values than the material formulated with EDAB, giving them high shelf-life stability. The degree of conversion after shelf simulation was only reduced for the EDAB material. Ultimate tensile strength, translucency parameter, and micro-tensile bond strength to dentin were similar for control and experimental adhesive resins. Due to their characteristics, TTME and TTAC monomers are potentially useful in the formulation of photopolymerizable resins for dental use with high shelf-life stability.This work reports the use of two monomers with two tertiary amines and four methacrylic (TTME) or acrylic (TTAC) terminal groups as co-initiators in the formulation of experimental resin adhesive systems. Both monomers were characterized by FT-IR and  Adhesive systems in dentistry are used to achieve the adhesion of restorative materials to dental structures . The useDental adhesive systems generally contain dimethacrylate monomers, organic solvents, and a photo-initiator system. The most used photo-initiator system contains camphorquinone (CQ) as a photo-initiator and ethyl-4-dimethylaminobenzoate (EDAB), a tertiary amine, as a co-initiator . CQ prodConsidering this, the objective of this study was the evaluation of the TTME and TTAC monomers in the formulation of experimental adhesive resins without EDAB. The use of TTME and TTAC results in similar properties to the control adhesive resin formulated with the conventional co-initiator (EDAB). Additionally, the experimental monomers have two amino groups and four terminal double bonds within their structure. The amino groups allow them to act as co-initiators, while the four double bonds guarantee that all of the initiator forms part of the adhesive resin, preventing its migration. In addition, the TTMEE and TTAC co-initiators cause a decrease in the polymerization rate, and therefore the experimental resins have high shelf-life stability.2. The irradiance was periodically monitored using a digital radiometer . Mechanical properties were evaluated using a universal Instron 4465 mechanical testing machine . Confocal microscopy images were obtained with a confocal laser scanning microscope . For each methodology, the sample size was estimated based on the data of previous studies, considering a comparative study design with three independent groups, a power of 0.8, and α = 0.05.The dimethacrylate monomers bisphenol A-glycidyl methacrylate (BisGMA), triethylene glycol dimethacrylate (TEGDMA), and (hydroxyethyl)methacrylate (HEMA), camphorquinone (CQ), and ethyl 4-(dimethylamino)benzoate (EDAB) used in the formulation of adhesive resins and 4,4′-methylenebis , methacrylic acid, and acrylic acid used in the synthesis of the monomers were purchased from Sigma-Aldrich . The FT-IR spectra were obtained with a Perkin Elmer Frontier spectrophotometer using the reflectance (ATR) technique . The solvents used were distilled with the techniques described in the literature. The NMR spectra were obtained with a 400 MHz spectrometer  using deuterated chloroform as a solvent and tetramethylsilane as an internal reference. The photopolymerization of the materials was carried out using a Bluephase N photopolymerization unit (Ivoclar-Vivadent) with an average irradiance of 1200 mW/cmThe TTMEE and TTAC were synthesized according to a methodology described elsewhere : 1.0 g . The rea−1): 2962 (νC-N), 1692 (νC=O), 1623 (νC=C), 1523 (νC=Caromatic). 1H-NMR : 7.05 , 6.72 , 6.20 , 6.20 , 5.6 , 3.70 , 3.40 , 3.15 , 2.75 , 2.55 , 1.92 .TTMEE characterization. FT-IR/ATR (cm−1): 2922 (νC-N), 1732 (νC=O), 1623 (νC=C), 1504 (νC=Caromatic). 1H-NMR : 7.01 , 6.67 , 6.44 , 6.07 , 6.20 , 5.88 , 4.06 , 3.64 , 3.34 , 3.12 , 2.76 , 2.62 .TTAC characterization. FT-IR/ATR  was determined according to the following equation 1/2, where w and b refer to the color coordinates measured on the white and black backgrounds.Composite-adhesive disks were then removed from the silicon mold and the CIELAB color parameters were measured using a spectrophotometer . All specimens were measured over white (equation :(3)ΔE00=Extracted human third molars were used after approval of the Ethical Review Board from the School of Medical Sciences at the Autonomous University of Hidalgo State (protocol CEEI-032-2019). Immediately after extraction, all blood and adhered tissue were removed, and thereafter stored in 0.5% chloramine/water solution at 4 °C for one week. Then, teeth were stored in distilled water at 4 °C for no longer than two months until their use . For specimen preparation, the root was sectioned, and the crowns were embedded in acrylic resin, allowing the buccal enamel surface to be exposed. Then, the occlusal part of the crowns was removed with an orthodontic grinder until exposure of a flat mid-coronal dentin surface. The exposed dentin surface was then wet-abraded using 600-grit silicon carbide sandpaper for 30 s to produce standardized smear-layers.TM Z250) were constructed in 3 increments of 2 mm each, and each layer was polymerized for 30 s. Light-curing procedures were performed using an LED photopolymerization unit, Bluephase N (Ivoclar-Vivadent). After immersion in distilled water at 37 °C for 24 h, the specimens were sectioned using a slow-speed diamond saw  to obtain resin-dentin sticks with a cross-sectional area of approximately 0.9 mm2. After storage in distilled water at 37 °C for 24 h, the sticks were individually fixed to a tensile testing device with cyanoacrylate glue and the TBS was tested in a mechanical universal test machine (Instron 4465), at a crosshead speed of 1 mm/min with a 1000 N load cell.Dentin specimens were randomly divided into three groups based on the adhesive system used (n = 5). Before applying the adhesive resins, a commercial self-etch primer  was applied according to the manufacturer’s instructions. Then, the corresponding adhesive resins were applied and rubbed for 10 s into the surface and photoactivated for 10 s. After the bonding procedures, resin composite build-ups  was added into the respective adhesives at a concentration of 0.08%. After the bonding procedures, one composite increment was placed over the adhesive resin. Then, the teeth were cut longitudinally into two halves and the generated surfaces were polished for one minute with SiC paper in sequence (Grit 800/1200/2000/4000) under water cooling. The samples were examined in 1000-fold magnification under a CLSM  at a 514 nm excitation line of the argon ion laser. The emissions were detected using a DD 458/514 band-pass filter.p = 0.05.Statistical analysis was performed using Sigma Plot 12.0 software. The data were analyzed to verify the normal distribution and homogeneity of the variance. A one-way analysis of variance (ANOVA) was used to compare the dependent variables. In all cases, the level of significance was set at 1H-NMR spectroscopies. FT-IR spectra of these compounds revealed that the absorption band at 910 cm−1, corresponding to the epoxy group of the initial compounds, disappears, suggesting the opening of the epoxy ring. The latter is confirmed by the appearance of the elongation vibration of the hydroxyl group at 3428 cm−1. In addition, new absorption bands located at 1711 cm−1, assigned to the elongation vibration mode of the carbonyl group νC=O, and at 1629 cm−1, assigned to the elongation vibration of the νC=C bond of the methacrylic groups, suggest the incorporation of the acryloyl and methacryloyl functional groups within the structure of the raw material .The sensitivity to ambient light of each adhesive is shown in p > 0.05), and in the case of the TTAC adhesive, the DE shift observed was statistically significantly lower (p < 0.05). Additionally, none of the adhesive systems tested in this study promoted statistically significant differences in the translucency of the composite (p = 0.893).p = 0.820). Confocal light scanning microscopy images of the bonded interfaces are shown in pmax values and similar degrees of conversion values compared to EDAB. After shelf-life simulation and sensitivity to ambient light, experimental adhesives showed better stability than the EDAB adhesive resin. With regards to UTS and bonding properties, they were similar to those presented by the EDAB adhesive. Considering this, the hypothesis was partially accepted.In this work, the synthesis of two tetramethacrylates, TTAC and TTME, was reported. They were used as co-initiators in the formulation of experimental adhesive resins. The monomers have two amino groups and four terminal double bonds within their structure. The amino groups allow them to act as co-initiators, while the four double bonds guarantee that all of the initiator forms part of the adhesive resin, preventing its migration. Their chemical–mechanical performance was compared with an adhesive resin formulated with EDAB, used as a control. Overall, TTAC and MBTTME achieved lower Rpmax values . The mechanical properties of polymers mostly depend on the adequate conversion of double bonds during polymerization [The mechanical properties of the adhesives were also evaluated. The mechanical property was measured in terms of ultimate tensile strength (UTS), a property that is frequently used to evaluate the performance of an adhesive in terms of its tensile bond strength with the local tissue . The resrization ; besidesrization . In thisp > 0.05), and in the case of the TTAC adhesive, the DE shift observed was lower (p < 0.05). Additionally, none of the adhesive systems tested in this study promoted statistically significant differences in the translucency of the composite (p = 0.893), so it can be suggested that the incorporation of the TTAC or TTME monomer into an experimental adhesive resin would not be clinically perceivable.Some optical parameters of the experimental adhesives were also evaluated . The newFinally, the micro-tensile bond strength (µ-TBS) to dentin and the failure mode of adhesive resins were tested using in vitro test methods that provide reliable data for materials’ development and/or evaluation of experimental variables. The micro-tensile bond strength test is currently recommended as the best method to evaluate the bond strength of adhesive systems, and it is considered useful for preliminary evaluation as a pre-clinical test . AccordiThe TTME and TTAC monomers were successfully used as polymerizable co-initiators in the formulation of experimental resin adhesive systems. The characteristics shown by the experimental materials can prove that the use of TTME and TTAC as polymerizable co-initiators represents an adequate alternative for the development of new resin-based materials. TTME and TTAC could provide high shelf-life stability and extended working time compared to the conventional materials formulated with EDAB.


The 44-year-old male healthcare worker who had frequent professional contacts with COVID-19 patients was hospitalized with a viral pneumonia diagnosis and soon started to exhibit fever, dry paroxysmal cough, loss of smell, and typical ground-glass opacities found in both lungs on chest CT scans. The COVID-19 diagnosis was verified by real-time quantitative polymerase chain reaction (qRT-PCR), immunochromatography, and immunochemiluminescent assays. The patient was treated with hydroxychloroquine, azithromycin, paracetamol, and enoxaparin, leading to his recovery after two weeks from the disease onset. The virus was successfully isolated from the nasopharyngeal swab sample taken on the fifth day of the disease onset using the Vero E6 cell line and exhibited a pronounced cytopathic effect (CPE) with a viral titer reaching 106 TCID50/ml in the cell culture medium. The full genome sequence of the viral isolate was obtained and 8 nucleotide and 5 amino acid mutations compared to the Wuhan-Hu-1 reference genome were identified. Viral isolate belonged to GR / 20B / B.1.1 genetic lineage  - the most prevalent genotype found in Russia to date.The coronavirus disease 2019 (COVID-19) pandemic is probably the most studied one in history from both clinical and molecular-epidemiological perspectives. Nonetheless, data on the correlation between severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) viral genotypes and COVID-19 symptoms caused by them are still scarce. In this report, we present a moderately severe COVID-19 case in a healthcare worker in Moscow, Russia, supplemented with the data on its causative agent's phenotype regarding  The eThe patient, a 44-year-old male, was hospitalized on 30.03.2020 (the second day after the onset of the disease) with viral pneumonia. Frequent professional contacts with COVID-19 patients were described in his medical history. Upon admission to the hospital, the patient’s condition was assessed as moderate with complaints of weakness, fatigue, and fever. Since the first day of the disease onset (29.03.2020), the patient had exhibited moderate fatigue and chills, but his temperature had not been measured and he had taken paracetamol to alleviate the symptoms. On the third day, a dry paroxysmal cough appeared, and on the fourth day, the patient noted the loss of smell. The fever persisted for 10 days with body temperature ranging from 37.4 to 39.2 °С. The patient was on eprosartan + hydrochlorothiazide 600 mg, atorvastatin 40 mg, and nebivolol 2.5 mg for his arterial hypertension.On examination, skin and sclera were of normal color, with no rash. The mucous membrane of the oropharynx was moderately hyperemic, tonsils were not enlarged, with no plaque. Peripheral lymph nodes were not enlarged and were painless on palpation. Breathing was not impaired; the frequency of respiratory movement was 18 per minute, with SpO2 of 96%. Heart sounds were rhythmic, without abnormal sounds, heart rate was 78 beats per minute, and blood pressure was 130/90 mmHg. The abdomen was soft and painless upon palpation, and the liver was at the costal margin. The consciousness was clear, with no focal or meningeal symptoms. Urination was not impaired, and there was no diarrhea.Blood tests and biochemical results in dynamic are presented in Tables Chest CT scans were performed on days two and eight of the disease onset with the following results: 30.03.2020 - up to 25% damage to both lungs with ground-glass opacities; 05.04.2020 - up to 45% damage with the progression of ground-glass opacities to infiltration Figure . Chest XThe COVID-19 diagnosis was verified by polymerase chain reaction (PCR), immunochromatography, and immunochemiluminescent assays Table . SARS-CoThe patient was given detoxification and symptomatic therapy, including hydroxychloroquine , azithromycin (500 mg once per day), paracetamol (up to 4 g per day with a fever above 38.5 °С), and enoxaparin 0.4 ml every 12 hours. On the ninth day of the disease, the patient recovered his sense of smell; his body temperature normalized on the next day, and he was discharged after 14 days from the onset of the disease.A PCR-positive nasopharyngeal swab (taken on the fifth day of the disease) was used to isolate the virus in a Vero E6 cell culture followed by viral genome high-throughput sequencing. The obtained viral isolate exhibited a pronounced cytopathic effect (CPE) on the third day of cell line inoculation Figure  with a vGenetic analysis showed 8 nucleotide and 5 amino acid substitutions between PMVL-6 and reference strain Wuhan-Hu-1 (GenBank accession NC_045512.2) in the following regions: 5’-untranslated region (5’-UTR), nsp3 (multi-domain transmembrane protein), nsp12 , nsp13 (helicase), S (spike glycoprotein) and N (nucleocapsid protein) . It includes SARS-CoV-2 isolates circulating in 93 different countries (as of 07.10.2020), the most represented of which are the United Kingdom , USA , and Australia . Of particular interest among detected amino acid substitutions is spike D614G mutation, which is present in almost all genomes of viral strains belonging to 20A, 20B, and 20C clades (Nextstrain nomenclature). This substitution is possibly linked with increased SARS-CoV-2 virulence due to its effect on the destabilization of S protein conformation [in vitro is crucial for further improvements in molecular diagnostics, antivirals, and vaccine development.The case we described is one of the very few globally published case reports of COVID-19 from Russia, and despite its typical clinical picture, it might be of interest to the global community, since it depicts infection with the most common Russian viral genotype to date. The possibility of SARS-CoV-2 nosocomial infections highlights the significance of stringent preventive measures in hospitals and our case highlights this problem, considering the most likely route of the infection in our patient. Continuous characterization of viral genomes and phenotypes


The aim of this study was to evaluate whether the portocaval shunt (PCS) corrects these unwanted changes in transhepatic flow after extended hepatectomy (EH). Forty female Landrace pigs were divided into two main groups: (A) EH (75%) and (B) no EH. Group A was divided into 3 subgroups: (A1) EH without PCS; (A2) EH with side-to-side PCS; and (A3) EH with end-to-side PCS. Group B was divided into 2 subgroups: (B1) side-to-side PCS and (B2) end-to-side PCS. HAF, PVF, and PVP were measured in each animal before and after the surgical procedure. EH increased the PVF/100 g  and PVP  but reduced the HAF/100 g . Following EH, side-to-side PCS reduced the increased PVF  and PVP . Without EH, side-to-side PCS reduced the PVF/100 g  and PVP . PVP was reduced by end-to-side PCS following EH by 48% (p < 0.001) and without EH by 21% (p = 0.075). PCS can decrease and correct the elevated PVP and PVF/100 g after EH to close to the normal values prior to resection. The decreased HAF/100 g in the remnant liver following EH is increased and corrected through PCS. Improvements in patient selection criteria, surgical methods, and postoperative care have increased the indications for therapeutic extended hepatectomy (EH)5. The most important complication of EH is posthepatectomy liver failure or small for size and flow (SFSF) syndrome, which is associated with significantly high rates of morbidity and mortality7. The current therapeutic methods are not effective because most often, early tissue damage following SFSF is irreversible, and the liver parenchyma loses its capability to regenerate. Thus, the best approach is to predict the chance of SFSF and to perform the appropriate preventive procedure.Liver resection is the most common and most efficient treatment for primary and secondary hepatic tumors, one that can provide a chance of long-term survival11. Portal vein flow (PVF), hepatic artery flow (HAF), and portal vein pressure (PVP) have been described as the main critical parameters for the development of SFSF13. Following EH, the ratio of HAF to the remnant liver weight (HAF/100 g) decreases, while PVF/100 g and PVP increase, resulting in various pathologic consequences that lead to SFSF16. Troisi et al. recommended a flow of 250 ml/min/100 g as the upper limit for PVF to prevent SFSF syndrome17. Thus, vascular modulation, which is able to decrease PVF/100 g and PVP, as well as increase HAF/100 g following EH, may prevent the occurrence of SFSF21. The aim of this study was to determine whether a portocaval shunt (PCS) can correct the unwanted changes that occur in transhepatic flow following EH.Current viewpoints strongly indicate that the incidence of SFSF after EH is directly dependent on transhepatic flow and remnant liver volumeForty female Landrace pigs aged between 10 and 12 weeks and weighing between 29 and 34 kg (mean 31 ± 2.7 kg) were included in this study. There was no significant difference between the groups with regard to the age or weight of the animals.Hemodynamic variables remained stable in all animals for the full duration of the experiment. The MAP, CVP, and HR showed maximum changes of 5 mmHg, 1 mmHg, and 14 beats/min before and after each procedure, respectively, which were not statistically significant. The mean blood loss volume was 75 ± 20 ml during the procedures.After EH, the PVF/100 g of remnant liver increased from 102.11 ± 9.09 to 279 ± 36.44 ml/min/100 g . This correction was also seen in HAF following EH through end-to-side PCS. As mentioned in our previous study, it is important to compare the results proportional to the remnant liver volume44. However, end-to-side PCS can be an option in few cases with chronic portal vein obstruction. Although short-term follow-up results are necessary and form a basis for further long-term studies, the absence of a long-term follow-up is admittedly a limitation of this study.PCS can be performed either side-to-side or end-to-side. Based on our results, the side-to-side PCS corrects the PVF and PVP changes following EH up to the normal values in an intact liver (without resection). The end-to-side PCS could show better results in regard to the HAF. This is because the liver loses the whole PVF following an end-to-side PCS and attempts to compensate for this variation by increasing the HAF. In the clinical setting, side-to-side PCS is more practical than end-to-side PCS. Redirecting the whole PVF through an end-to-side PCS can result in high-stage hepatic encephalopathy as well as right heart failureHepatic inflow modulation by PCS following EH is feasible. PCS following EH immediately reduces and corrects the increased PVF and PVP and increases the decreased HAF. Based on these findings and since the changes in HAF, PVF, and PVP are considered triggers of SFSF syndrome, further long-term experimental and clinical studies must be performed to evaluate the effectiveness of PCS in preventing SFSF syndrome following EH.45. This experimental study was conducted on forty female Landrace pigs. The animals were divided into two main groups: (A) EH (75%) and (B) no EH. Group A was further divided into 3 subgroups: (A1) EH without PCS; (A2) EH with side-to-side PCS; and (A3) EH with end-to-side PCS. Group B was divided into 2 subgroups: (B1) side-to-side PCS without EH and (B2) end-to-side PCS without EH 37. A Lohmeier M111 device  was used to monitor PVP, HAF, and PVF, as described in our previous study25. After performing any intervention, we monitored the PVF, HAF, and PVP in each study group until the values reached a stable level. The HAF and PVF are reported in ml per 100 g remnant liver.After a 12-h fasting time with free access to water, the pigs were anesthetized following our standardized narcotic protocol. Two catheters were placed in the carotid artery and internal jugular vein to monitor MAP and CVP, respectively. The heart rate, body temperature, and blood oxygen saturation were monitored during the entire procedure46. After EH, complete hemostasis was achieved by electrocoagulation and hand suturing. To perform the PCS, the portal vein and vena cava were prepared and released from the surrounding tissues in the potential anastomosis place. Both veins were cross-clamped after heparinization with 30 IU/kg. For side-to-side PCS placement, a portal vein-caliber size anastomosis was performed between the right lateral side of the portal vein and medial side of the vena cava using Prolene 5–0  in a continuous technique . In accordance with the institutional guidelines established for the Animal Care Facility at the University of Heidelberg, all animals received human care during the experiment. Once the surgical procedures were completed and the hemodynamic measurements were obtained, the animals were euthanized by an intravenous injection of potassium chloride (2 mmol/kg) under deep anesthesia.The statistical analysis was performed using SPSS 22 . All measured values are expressed as the mean ± standard deviation (SD), and the changes in the values are graphically represented. The differences in mean hemodynamic measurements between study groups were tested using independent samples t tests. A p value less than 0.05 was considered to be statistically significant in all tests.The study protocol was approved by the German Committee for Animal Care, Karlsruhe, Germany (AZ: 35–9185.81/G-45/12).


Background: Primary Sjögren’s syndrome (pSS) is a chronic systemic autoimmune disease characterized by typical autoantibody production and lymphocytic-mediated exocrine gland damage. Interstitial lung disease (ILD) is a common complication of pSS and can be associated with a poor prognosis. However, the pathogenesis of ILD in pSS is still unclear.Methods: In this study, we used RNA sequencing to investigate the gene-expression profile of the minor salivary glands (MSGs) from 36 patients with ILD-pSS and 128 patients with non-ILD-pSS.Results: In the remarkably enriched chemokine-mediated signaling pathway, C-X-C motif chemokine receptor 2 (CXCR2), a receptor for interleukin-8, which participates in the activation of neutrophils, was found to be significantly elevated in both MSG and plasma from pSS patients with vs. without ILD (p < 0.001). Furthermore, the CXCR2 expression level in MSG and plasma was significantly associated with the diffusing capacity of the lungs for carbon monoxide, erythrocyte sedimentation rate, and EULAR Sjögren’s Syndrome disease Activity Index in ILD-pSS.Conclusion: Therefore, with its potential role in ILD progression in patients with pSS and its strong association with clinical manifestations of the disease, CXCR2 may serve as a useful index for disease activity in ILD associated with pSS. Primary Sjögren’s syndrome (pSS) is a systemic autoimmune disease characterized by the presence of organ-specific or non-specific autoantibodies and lymphocyte-mediated exocrine gland damage . Althoug2), are considered to be of great significance for pSS  biopsy is a cornerstone in the diagnosis of pSS . MSG bio for pSS . Further for pSS . Gene-ex for pSS . HoweverIn this study, we investigated the clinical characteristics of and compared gene expression in MSGs from patients with ILD-pSS and non-ILD-pSS, with the aim of enhancing our understanding of the mechanisms underlying the progression of patients with pSS with ILD. In addition, we explored potential biomarkers for evaluating the disease severity of ILD-pSS in clinical practice.n = 36) and non-ILD-pSS (n = 128) were recruited from the First Affiliated Hospital of Wenzhou Medical University, China, between 1 January 2018 and 30 June 2020. All patients fulfilled the 2016 American College of Rheumatology (ACR)/EULAR classification criteria  scores were obtained before the participants were given any systemic immunosuppressant or glucocorticoid to avoid confounding effects of these medications.The study was approved by the ethics committee of the First Affiliated Hospital of Wenzhou Medical University , and all patients gave their written informed consent.Total RNA was isolated from frozen salivary gland samples using TRIzol Reagent  according to the manufacturer’s instructions. RNA purity was checked using a Nano Photometer spectrophotometer . The quality of the RNA used to construct the cDNA library was verified (RNA integrity numbers >9) using an RNA 6000 Nano kit with the Bioanalyzer 2100 system . With input material of 3 mg RNA per sample, we constructed sequencing libraries using the NEBNext Ultra RNA Library Prep Kit for Illumina  and subsequently performed the sequence on an Illumina HiSeq platform. Then, using Cutadapt adapters and TrimGalore, raw reads were trimmed and low-quality reads were filtered. Quality-control reports of sequence reads were obtained using FastQC software. Finally, the sequencing data were aligned to the human reference genome “hg38” using STAR software, and the read count files were filtered with low expression and normalized using the DESeq2 package.Levels of C-X-C motif chemokine receptor 2 (CXCR2) and interleukin (IL)-8 in the plasma of patients with ILD-pSS and non-ILD-pSS were quantified using Human CXCR2 and Human IL-8 ELISA kits , respectively according to the manufacturer’s instructions. The color was developed using 3,3′,5,5′-tetramethylbenzidine  and then measured using an ELISA plate reader . All samples were measured in triplicate.t-test was performed, calculating the mean ± standard error (SE). Differentially expressed genes (DEGs) were identified according to the following criteria: adjusted p value < 0.05 and absolute value of log2 fold change >1. Pearson’s correlation analysis was used to quantify the associations between gene-expression and clinical parameters, and Wilcoxon’s test was applied to comparisons between subgroups. p values <0.05 were considered significant.The data analysis was conducted using R version 3.6.1. Two-sample The characteristics of the 36 patients with ILD-pSS and the 128 patients with non-ILD-pSS are detailed in p < 0.001; p < 0.001; p = 0.059; p = 0.097) or anti-SSB antibodies (p = 0.144), or in the levels of immunoglobulin (Ig)G, IgA, IgM, C3, or C4  ESSDAI scores were elevated in those with ILD-pSS (20.64 ± 9.37) and significantly higher than in patients with non-ILD-pSS . In addition, CXCR2 was of greatest interest because of its potential role in inflammation . Concentrations of CXCR2 in plasma were positively correlated with the ESSDAI score (r = 0.59, p < 0.001; r = 0.59, p < 0.001; r = –0.30, p = 0.08; Concentrations of CXCR2 and IL-8 in plasma were detected by ELISA in patients with ILD-pSS and non-ILD-pSS. CXCR2 levels were higher in those with ILD-pSS (CXCR2. Moreover, the CXCR2 level correlated with clinical features in these patients, implying a potential role for CXCR2 in the progression of ILD in pSS.In this study, we first compared the MSG gene-expression profiles of 36 patients with ILD-pSS and 128 patients with non-ILD-pSS, and identified several genes that are up-regulated in ILD-pSS. To identify potential biomarkers for evaluating disease severity in ILD-pSS, the clinical implications of the top 20 DEGs were examined. In particular, we found that patients with ILD-pSS had abnormally increased levels of CXCR2 (CXCR2 is expressed by granulocytes, especially neutrophils (CXCR2-mediated neutrophil recruitment is essential for lung fibrosis (CXCR2 can reduce lung fibrosis (CXCR2 was found to be elevated in both the MSG and plasma of patients with ILD-pSS vs. non-ILD-pSS, with the results showing that CXCR2 may also play an essential part in organ-typical damage and circular inflammation in pSS. Although patients with ILD-pSS and those with non-ILD-pSS may initially show similar clinical manifestations (CXCR2  is a G-ptrophils . Neutropfibrosis , and thafibrosis . In thisstations , those wstations . FurtherCXCR2 in both MSG and plasma was highly associated with the ESR, indicating that CXCR2 may help differentiate ILD-pSS from non-ILD-pSS in some situations. The ESSDAI was developed to measure systemic disease activity in pSS (CXCR2 levels in both MSG and plasma from patients with ILD-pSS, indicating that CXCR2 may serve as a useful candidate for evaluating disease severity. DLCO is one measure for predicting lung function. Most previous studies (CXCR2 levels in both MSG and plasma from patients with ILD-pSS were inversely associated with DLCO, confirming the role of CXCR2 in evaluating the disease severity of ILD in those with pSS.The ESR is a common inflammatory marker, and an elevated ESR is found associated with ILD underlying other connective tissue diseases . Furthery in pSS . In our  studies  used a sCXCR2 in ILD-pSS. We observed extensive expression of CXCR2 in both MSG and plasma in patients with ILD-pSS, and many manifestations of ILD are characterized by the accumulation of inflammatory cells. CXCR2 is implicated in neutrophil infiltration and can be activated by many other CXC chemokines in inflammatory process. Therefore, the contribution of CXCR2 to the inflammatory and further fibrotic process in ILD-pSS should be further investigated.To our knowledge, this is the first study to suggest a possible role for CXCR2 in patients with ILD-pSS. CXCR2 may serve as a potential biomarker for evaluating disease severity in this population.Our results provide new insights into the role of


Pakistan has not been a major contributor to medical research, mainly because of the lack of learning opportunities to medical students. With the increase in online learning systems during COVID-19, research related skills can be taught to medical students via low-cost peer taught virtual research workshops.To assess the effectiveness of a comprehensive low-cost peer-taught virtual research workshops amongst medical students in Pakistan.This quasi-experimental study assessed the effectiveness of five virtual research workshops (RWs) in improving core research skills. RWs for medical students from across Pakistan were conducted over Zoom by medical students (peer-teachers) at the Aga Khan University, Pakistan, with minimal associated costs. The content of the workshops included types of research, ethical approval and research protocols, data collection and analysis, manuscript writing, and improving networking skills for research. Improvement was assessed via pre-and post-quizzes for each RW, self-efficacy scores across 16 domains, and feedback forms. Minimum criteria for completion of the RW series was attending at least 4/5 RWs and filling the post-RW series feedback form. A 6-month post-RW series follow-up survey was also emailed to the participants.p <  0.001) with the greatest improvement in Data Collection and Analysis (+ 34.65%), and in self-efficacy scores across all domains (p <  0.001). 166/307 (54.1%) participants responded to the 6 months post-RWs follow-up survey. Compared to pre-RWs, Research involvement increased from 40.4 to 62.8% (p <  0.001) while proportion of participants with peer-reviewed publications increased from 8.4 to 15.8% (p = 0.043).Four hundred medical students from 36  different medical colleges in Pakistan were enrolled in the RWs. However, only 307/400 (76.75%) medical students met the minimum requirement for completion of the RW series. 56.4% of the participants belonged to the pre-clinical years while the rest were currently to clinical years. The cohort demonstrated significant improvement in pre-and post-quiz scores for all 5 RWs (Virtual RWs allow for a wide outreach while effectively improving research-related knowledge and skills, with minimal associated costs. In lower-middle-income countries, virtual RWs are a creative and cost-effective use of web-based technologies to facilitate medical students to contribute to the local and global healthcare research community. Health research is crucial for practicing evidence-based medicine . PhysiciMoreover, only few medical students from low-middle-income countries (LMICs) like Pakistan are involved in research . The parEarly exposure to research can be beneficial for medical students in improving skills such as searching and critically appraising the medical literature, independent learning, and manuscript writing . MoreoveThe COVID-19 pandemic has created a health care crisis around the world and has presented a spectrum of challenges, including a profound effect on the delivery of education. Institutions are being forced to transition to online and remote education . Moreoveth and 26th July 2020 over the online video conferencing platform Zoom, due to feasibility during the ongoing COVID-19 pandemic. The technical analysts’ team at CIME, AKU provided the technical support and service required for the RWs The peer-taught research workshops (RWs) were carried out after prior approval from the institutional ethics review committee .A quasi-experimental study was conducted by the Society for Promoting Innovation in Education (SPIE), in collaboration with the International Federation of Medical Students’ Associations (IFMSA) – Pakistan, Department of Biological and Biomedical Sciences (BBS) and the Center for Innovation in Medical Education (CIME), at the Aga Khan University (AKU), Pakistan. Five research workshops (RWs) were conducted between 20In Pakistan, medical school is a 5-year undergraduate program  that is entered directly after high-school i.e., enrolment in a pre-medical college program is not a pre-requisite. In medical schools in Pakistan, the first 2 years (Years 1 and 2) are pre-clinical years, where students are taught the basic sciences. Following that, Years 3–5 are clinical years, during which students complete clinical rotations. Throughout medical school, a variety of teaching pedagogies are employed, including conventional didactic lectures, problem-based learning, and team-based learning. The degree to which these different pedagogies are used amongst medical schools.The Society for Promoting Innovation in Education (SPIE) is a student-run, non-profit, organization, founded in 2017 by the medical students at AKU. SPIE aims to promote culture of innovative learning and revolutionize educational practices. SPIE has five wings, including Research and Development Wing (R&D Wing). The RWs were organized by SPIE’s R&D Wing, in collaboration with International Federation of Medical Students Associations (IFMSA), Pakistan. Through this collaboration, SPIE was able to expand its outreach to medical colleges across the country, both public- and private-sector, hence improving national generalizability. Additionally, SPIE has previously published a protocol  for a siThe minimum required sample size calculated using University of California San Francisco Calculator  was 161,n = 117) [n = 29), Punjab (n = 62), Baluchistan (n = 2), Khyber Pakhtunkhwa (n = 20), and Azad Jammu and Kashmir (n = 4).The recruitment for participants of the workshops was carried out via a Google Form which was disseminated on SPIE’s social media platforms including Facebook, Twitter and WhatsApp. The target population of the RW participants consisted of medical students from all over Pakistan. The inclusion criteria consisted of students aged 18 years or above currently enrolled in an MBBS program in a Medical College of Pakistan. The first 400 registrations meeting the inclusion criteria were selected and enrolled (convenience sampling). Despite the minimum required sample size being 161 only, 400 participants were enrolled to maximize benefit of the RW series at no additional cost or compromise in the quality of the RWs. A confirmation email, along with a consent form see  was sentn = 117) : Sindh . In addition, students were asked if they had ever been involved in conducting research before, and if they had ever attended a RW before. Though all 400 participants would have filled the sign-up form, only those meeting minimum criteria for completion were included in the final analysis.Pre-and Post-RW Quizzes: Each participant was required to take a quiz before and after attending each RW, to evaluate the effectiveness of the respective RWs. Both the pre- and the post-quiz for each RW were identical, and tested participants’ research-related knowledge and application of research-skills. Out of the 400 enrolled participants, only those attending each individual RW were allowed to attempt the quiz for that specific RW. Students were only required to attempt the quizzes in order to fulfil attendance criteria, with scores on pre- and post-RW quizzes not impacting completion of the RW series. The pre- and post-RW quizzes were piloted during the aforementioned mock sessions, and ambiguous questions were amended. The Cronbach’s alpha for the pre- and post-RW quizzes were 0.61 and 0.78, respectively, indicating acceptable to good reliability of the quizzes.Pre- and Post- Self-Efficacy Forms: Participants were required to rate their confidence in 16 pre-defined objectives  was provided to all participants in the initial enrolment email, and all data collected subsequently was done so using the UIN. Only two research team members, RDU and MAB, assigned and emailed the UINs to selected participants, and they were also responsible for tallying attendance in order to provide Certificates of Completion. However, RDU and MAB did not partake in data handling, so as to maintain participants’ anonymity.p-value < 0.05 was regarded as significant for all analyses.Analysis was performed using IBM SPSS for Windows, version 23.0 . Categorical data was described as frequencies (n) and percentages (%). Continuous data was expressed as mean and standard deviation. Paired t-tests were used to compare mean pre-and post-research workshop quiz scores and self-efficacy scores. Categorical data was compared using Chi-squared tests, while continuous data was compared using independent sample t-tests. McNemar’s test was used to compare paired responses to dichotomous variables in the 6-month follow-up surveys . A Out of the 400 medical students enrolled from 36 medical colleges in Pakistan, a total of 307 (76.8%) medical students met the minimum completion requirements. Amongst the 36 medical colleges included, 21 (58.3%) were public-owned while the rest were private-owned. The province−/region-wise distribution of the 36 medical colleges were as follows: 17  from Punjab, 14  from Sindh, 3  from Khyber Pakhtunkhwa, 1  from Baluchistan, and 1  from Azad Jammu and Kashmir. The mean age of the students was 21.37 ± 1.91 years. Across the 5 RWs, the maximum attendance was seen in RW1  and RW3 , while lower attendance was seen in RW2 , RW4 , and RW5 . 56.4% of the participants belonged to the pre-clinical years, with a year-wise breakdown as follows: 80/307 (26.1%) from Year 1, 93/307 (30.3%) from Year 2, 26/307 (8.5%) from Year 3, 64/307 (20.8%) from Year 4, and 44/307 (14.3%) from Year 5. Most students (60.9%) did not have any prior research experience and the majority (59.0%) had not attended any Research workshop previously. The demographics are summarized in Table The students demonstrated significant improvement in scores from pre-workshop quiz to post-workshop quiz (p <  0.001). The greatest improvement in scores was shown in RW 3  with mean difference of 34.65% ± 24.49%. The results for pre-and post-workshop quizzes are summarized in Table p-value = 0.024). In addition, the difference in the mean improvement of pre-workshop and post-workshop quiz for RW 5 (Improving Networking Skills for Research) was significantly higher among the participants who had conducted research before compared to the participants who had not conducted research previously .The difference in the mean improvement of pre-workshop and post-workshop quiz for RW 5 (Improving Networking Skills for Research) was significantly higher in the pre-clinical students compared to clinical students . The greatest improvement in self-efficacy scores were for “Understanding an ANOVA test” (4.56 ± 3.33) followed closely by “Understanding the types of T-tests” (4.35 ± 3.33). The results are summarized in Table The pre- and post-workshop analysis revealed a significant improvement in all the 16 self-efficacy scores grouped into 5 categories (” 4.56 ± .33 follop = 0.006), “Understand Chi-Square tests” , Compared to participants without prior research experience, participants with prior experience demonstrated a significantly greater improvement in “Understanding an ANOVA test” .In comparison to participants with no prior research experience, the difference in the mean improvement of self-efficacy scores in “Understand the types of T-tests” .In comparison to the pre-clinical year students (79.2%), clinical years students believed that the time allotment was “Excellent/Good”  reported that participation in the RWs had enhanced their knowledge and skills to conduct research independently “Very Significantly/Significantly”, while the remainder reported that it had helped them “Moderately”. 62.8% students reported involvement in research projects during the 6-months after attending the RWs, as compared to their involvement before attending the RWs . In comparison to 14 (8.4%) participants with peer-reviewed publications before the RWs, 26 (15.8%) students reported having peer-reviewed publications 6 months after RWs (p = 0.043). The results of the follow-up survey are shown in Table Out of the 307 medical students who participated in the RWs, 166 participants responded to the 6-month follow-up survey (response rate: 54.1%). The majority (For the in-person RWs conducted by SPIE in January 2020, the total cost amounted to approximately USD 2235.36. The virtual RWs were conducted on Zoom. The technical analysts’ team at CIME, AKU provided the technical support and service required for the RW series. The Zoom package, already purchased by CIME, and technical support staff, was provided by CIME to SPIE for free of cost. There was no cost invested by SPIE. However, to present a holistic comparison of costs, support staff and Zoom costs are considered as expenditures for the virtual RWs. Even so, the virtual RWs catered to more than three times as many students as the in-person RWs at about 5% the total cost (Table The rapidly evolving crisis, the COVID-19 pandemic, has led to several challenges with respect to education. It has led to development of various forms of educational systems including peer teaching and utilization of virtual platform to meet the educational needs . The useResearch training is an important aspect of medical education to practice evidence-based medicine in healthcare . PerhapsIn our study, students demonstrated significant improvement in scores from pre-workshop quiz to post-workshop quiz and all the 16 self-efficacy scores pertinent to the basic research skills. Similarly, a study conducted by Morales-Pérez et al. reported significant improvement in the pre-and post- test scores of the participants after attending an online course . MoreoveVirtual mode of learning is more convenient, saves resources, quick and efficient, and gives students the opportunity to become independent learners . Our RW Peer teaching is an effective method of education delivery that promotes a learning environment among peers . The chaThis study had a few limitations. Though the pre-post design of this quasi-experimental study ensured a high internal validity, a degree of external validity is sacrificed due to the nature of the study design. Also, performance on the pre- and post-quizzes could be affected by external influences other than the RWs themselves, especially if participants attempted to supplement their learning at the RW series by consulting other resources. Additionally, there is no absolute way of knowing to what extent the heartening results at the 6-month follow-up in our study are attributable purely to the RW series.The pre-RW quizzes may have sensitized participants and led to bias in participants’ responses when answering the post-RW quizzes. In addition, there was no way invigilate students while they responded to the pre- and post-RW quizzes, and results may be biased by the honesty of participants’ quiz-taking behavior. Moreover, as scores on the quizzes did not impact workshop completion, participants may have not filled it with their complete attention. Furthermore, although self-efficacy has been used previously to evaluate learners’ perceptions of their own improvement, this rating system is undeniably subjective and does not in itself prove objective increase in expertise. Lastly, the lack of availability of internet facility in remote areas or unscheduled power cuts during the sessions were another challenge in the setting of an LMIC like Pakistan.The present study conducted by SPIE demonstrates the utility of virtual research workshops in improving the research-related knowledge and skills among medical students. The virtual nature of research workshops allows for a much wider outreach than conventional in-person research seminars. In addition, low-cost virtual research workshops are a creative use of web-based technologies to facilitate medical students to contribute to the local and global healthcare research community. Therefore, workshops conducted by SPIE can be easily replicated on a much larger scale to promote research culture in a cost-effective way.